Purification and Characterization of a Trypsin Inhibitor from Plathymenia foliolosa Seeds

• Published in: Journal of agricultural and food chemistry Vol. 56; no. 23; pp. 11348 - 11355
• Main Authors: da Silveira Ramos, Vanessa, de Souza Silva, Gilvânia, das Graças Machado Freire, Maria, Lima Tavares Machado, Olga, Postali Parra, José Roberto, Rodrigues Macedo, Maria Lígia
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 10.12.2008
• Subjects: Amino Acid Sequence; Anticarsia gemmatalis; Biological and medical sciences; chymotrypsin; Crop and Animal Protection Chemistry; Diatraea saccharalis; enzyme inhibition; Fabaceae - chemistry; Fabaceae - genetics; Fabaceae - metabolism; Food industries; Fundamental and applied biological sciences. Psychology; gel electrophoresis; insect pests; isolation; Kinetics; Kunitz family; larvae; midgut; Molecular Sequence Data; N-terminal sequence; pest protease; plant proteins; Plant Proteins - chemistry; Plant Proteins - genetics; Plant Proteins - isolation & purification; Plant Proteins - metabolism; Plathymenia; Plathymenia foliolosa; seeds; Seeds - chemistry; Seeds - genetics; Seeds - metabolism; Sequence Alignment; trypsin; trypsin inhibitor; trypsin inhibitors; Trypsin Inhibitors - chemistry; Trypsin Inhibitors - genetics; Trypsin Inhibitors - isolation & purification; Trypsin Inhibitors - metabolism; Kunitz family; Plathymenia foliolosa; trypsin inhibitor; N-terminal sequence; pest protease; Seeds; Purification; Serine endopeptidases; Enzyme; Characterization; Peptidases; Trypsin; Pest; N terminal-Sequence; Hydrolases; Inhibitor
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/jf802778b

A novel trypsin inhibitor (PFTI) was isolated from Plathymenia foliolosa (Benth.) seeds by gel filtration chromatography on a Sephadex G-100, DEAE-Sepharose, and trypsin-Sepharose columns. By SDS-PAGE, PFTI yielded a single band with a M r of 19 kDa. PFTI inhibited bovine trypsin and bovine chymotrypsin with equilibrium dissociation constants (K i) of 4 × 10−8 and 1.4 × 10−6 M, respectively. PFTI retained more than 50% of activity at up to 50 °C for 30 min, but there were 80 and 100% losses of activity at 60 and 70 °C, respectively. DTT affected the activity or stability of PFTI. The N-terminal amino acid sequence of PFTI showed a high degree of homology with various members of the Kunitz family of inhibitors. Anagasta kuehniella is found worldwide; this insect attacks stored grains and products of rice, oat, rye, corn, and wheat. The velvet bean caterpillar (Anticarsia gemmatalis) is considered the main defoliator pest of soybean in Brazil. Diatraea saccharalis, the sugar cane borer, is the major pest of sugar cane crops, and its caterpillar-feeding behavior, inside the stems, hampers control. PFTI showed significant inhibitory activity against trypsin-like proteases present in the larval midguts on A. kuehniella and D. saccharalis and could suppress the growth of larvae.